High resolution surface profilometry plays an important role in nanoscience and biological research. For instance, atomic force microscopy (AFM) can be used to study biomolecular activities in real time. Direct optical imaging techniques are also useful in biological research because of their capabilities in wide-field imaging and non-contact measurement. However, the lateral resolution of optical imaging systems is generally limited by diffraction to about 0.5λ, where λ is the wavelength of the imaging light.
Structured illumination microscopy (SIM) is an optical imaging technique with improved lateral resolution. In SIM, a periodic excitation pattern is projected onto a sample. Multiple images are taken with the modulating pattern shifted to different positions transversely to the optical axis. The modulated illumination patterns are eliminated from the collected images by the mathematical combination of multiple images, enabling the recovery of high spatial frequency information and the formation of high resolution images. Linear SIM provides a lateral resolution of about 0.25λ for fluorescence microscopy; even higher resolution is achievable with saturated SIM. Fluorescence SIM with axially sectioning ability also provides a resolution of about 0.25λ for one-dimensional or two-dimensional structured imaging.
Fluorescence SIM using a liquid-crystal spatial light modulator (SLM) simplifies the configuration of the microscopy system and improves the stability and imaging speed of the system. In one SIM algorithm, five images are used, with the modulating mesh pattern at a different position in each image, to obtain a fluorescence image with a lateral resolution of 0.3λ and an axial resolution of 0.38λ.